PROJECT SUMMARY Stress Pathways in the Induction of Metaplasia Mature cells in many organ systems can undergo a reprogramming event in which cells dedifferentiate, convert to a progenitor-like state, and enter the cell cycle in response to tissue injury. This cell state change results in the formation of metaplasia to repair lost and damaged tissue. If tissue is unable to return to normal after metaplasia formation this is a risk factor for cancer formation, as is the case with spasmolytic polypeptide- expressing metaplasia in the stomach and acinar-to-ductal metaplasia in the pancreas. The cellular and molecular mechanisms that govern injury-induced metaplasia formation are largely undescribed, yet they appear to occur in a consistent, step-wise manner: 1. Autodegradation of differentiated proteins, 2. Expression of a progenitor/metaplastic gene network, and 3. Entry to the cell cycle. A recent publication from our lab demonstrated that these steps are conserved in mouse injury models in the stomach, pancreas, liver, and kidneys. The experiments presented in this proposal aim to interrogate the importance of two proteins known to be involved in the integrated stress response, eukaryotic initiation factor 2 (eIF2) and activating transcription factor 3 (ATF3), during the development of metaplasia. Preliminary results show that eIF2 becomes inactivated rapidly after injury in mouse models that induce pancreatic and gastric metaplasia. ATF3 protein and Atf3 mRNA also rapidly increase in these injury models concurrent with eIF2 inactivation. The Aims presented here will further determine if Atf3 function and eIF2 inactivation are necessary components of metaplasia initiation with detailed mechanistic studies using Atf3-null mice, pharmacological inhibitors of eIF2, and an injury model in Caco-2 cell lines. Experiments will also determine how eIF2 becomes inactivated in this injury response and identify genes differentially expressed by ATF3. If the experiments presented here support the hypothesis that eIF2 and ATF3 are critical for mature cell reprogramming following injury, future studies will determine the importance of eIF2 and ATF3 in human gastric and pancreatic metaplasia and adenocarcinoma.